Transgenerational interactions between pesticide exposure and warming in a vector mosquito

While transgenerational plasticity may buffer ectotherms to warming and pesticides separately, it remains unknown how combined exposure to warming and pesticides in the parental generation shapes the vulnerability to these stressors in the offspring. We studied the transgenerational effects of single and combined exposure to warming (4°C increase) and the pesticide chlorpyrifos on life history traits of the vector mosquito Culex pipiens. Parental exposure to a single stressor, either warming or the pesticide, had negative effects on the offspring: both parental exposure to warming and to the pesticide resulted in an overall lower offspring survival, and a delayed offspring metamorphosis. Parental exposure to a single stressor did, however, not alter the vulnerability of the offspring to the same stressor in terms of survival. Parental pesticide exposure resulted in larger offspring when the offspring experienced the same stressor as the parents. Within both the parental and offspring generations, warming made the pesticide more toxic in terms of survival. Yet, this synergism disappeared in the offspring of parents exposed to both stressors simultaneously because in this condition the pesticide was already more lethal at the lower temperature. Our results indicate that transgenerational effects will not increase the ability of this vector species to deal with pesticides in a warming world. Bifactorial transgenerational experiments are crucial to understand the combined impact of warming and pesticides across generations, hence to assess the efficacy of vector control in a warming world.status: publishe

Evolution determines how global warming and pesticide exposure will shape predator‐prey interactions with vector mosquitoes

How evolution may mitigate the effects of global warming and pesticide exposure on predator–prey interactions is directly relevant for vector control. Using a space‐for‐time substitution approach, we addressed how 4°C warming and exposure to the pesticide endosulfan shape the predation on Culex pipiens mosquitoes by damselfly predators from replicated low‐ and high‐latitude populations. Although warming was only lethal for the mosquitoes, it reduced predation rates on these prey. Possibly, under warming escape speeds of the mosquitoes increased more than the attack efficiency of the predators. Endosulfan imposed mortality and induced behavioral changes (including increased filtering and thrashing and a positional shift away from the bottom) in mosquito larvae. Although the pesticide was only lethal for the mosquitoes, it reduced predation rates by the low‐latitude predators. This can be explained by the combination of the evolution of a faster life history and associated higher vulnerabilities to the pesticide (in terms of growth rate and lowered foraging activity) in the low‐latitude predators and pesticide‐induced survival selection in the mosquitoes. Our results suggest that predation rates on mosquitoes at the high latitude will be reduced under warming unless predators evolve toward the current low‐latitude phenotype or low‐latitude predators move poleward

Evolution of pesticide tolerance and associated changes in the microbiome in the water flea Daphnia magna

Exposure to pesticides can have detrimental effects on aquatic communities of non-target species. Populations can evolve tolerance to pesticides which may rescue them from extinction. However, the evolution of tolerance does not always occur and insights in the underlying mechanisms are scarce. One understudied mechanism to obtain pesticide tolerance in hosts are shifts toward pesticide-degrading bacteria in their microbiome. We carried out experimental evolution trials where replicated experimental populations of the water flea Daphnia magna were exposed to the pesticide chlorpyrifos or a solvent control, after which we performed acute toxicity assays to evaluate the evolution of chlorpyrifos tolerance. Additionally, we quantified changes in the microbiota community composition of whole body and gut samples to assess which sample type best reflected the pesticide tolerance of the Daphnia host. As expected, chlorpyrifos-selected clones became more tolerant to chlorpyrifos as shown by the higher EC50 48 h (36% higher) compared with the control clones. This was associated with shifts in the microbiome composition whereby the abundance of known organophosphate-degrading bacterial genera increased on average ~4 times in the chlorpyrifos-selected clones. Moreover, the abundances of several genera, including the organophosphate-degrading bacteria Pseudomonas, Flavobacterium and Bacillus, were positively correlated with the EC50 48 h of the host populations. These shifts in bacterial genera were similar in magnitude in whole body and gut samples, yet the total abundance of organophosphate-degrading bacteria was ~6 times higher in the whole body samples, suggesting that the gut is not the only body part where pesticide degradation by the microbiome occurs. Our results indicate that the microbiome is an important mediator of the development of tolerance to pesticides in Daphnia

Non-pathogenic aquatic bacteria activate the immune system and increase predation risk in damselfly larvae

1. Pathogens can increase vulnerability to predation through their harmful effects on hosts. Recently it was shown that the mere activation of the immune system by pathogens may increase the host’s risk of predation. Here we test whether exposure to non-pathogenic bacteria also activates the immune system and thereby increases vulnerability to predation. 2. We exposed Enallagma cyathigerum damselfly larvae to a non-pathogenic strain of the bacterium Escherichia coli and measured immune defence, anti-predator behaviour and survival times in the presence of larval dragonfly predators. To evaluate whether non-pathogenic bacteria also generated energy-based trade-offs leading to other fitness costs, we also quantified growth rate and survival in the absence of predators. 3. Exposure to the non-pathogenic bacterium did not affect survival in the absence of the predator but increased growth rate, possibly a response to reduce exposure time to the bacterium. Larvae exposed to the bacterium activated their immune response as shown by an increase in the activity of phenoloxidase and the number of haemocytes. The bacterium affected anti-predator traits involved in avoiding detection by predators as well as traits involved in escape after detection. Pre-exposed larvae showed higher activity levels and further increased the number of feeding strikes in the presence of predation risk, possibly driven by energetic constraints. Pre-exposed larvae swam less often when attacked, but faster. This impaired anti-predator response came at the ecological cost of increased vulnerability to predation. 4. Our study demonstrated that exposure to non-pathogenic bacteria increases vulnerability to predation, which is a novel type of antagonistic interaction. This highlights the unexplored possibility that non-pathogens may play a role in maintaining variation in immune defence through insidious effects on predator-prey interactions. Since non-pathogenic bacteria can be very abundant, this unexplored ecological cost of immune system activation in terms of increased predation may have major consequences in natural systems and may provide an unexplored new force underlying variation in immune defence.status: publishe

Reinforcing effects of non-pathogenic bacteria and predation risk: from physiology to life history

The important ecological role of predation risk in shaping populations, communities and ecosystems is becoming increasingly clear. In this context, synergistic effects between predation risk and other natural stressors on prey organisms are gaining attention. Although non-pathogenic bacteria can be widespread in aquatic ecosystems their role in mediating effects of predation risk has been ignored. We here address the hypothesis that non-pathogenic bacteria may reinforce the negative effects of predation risk in larvae of the damselfly Coenagrion puella. We found synergistic effects for all three life history variables studied: mortality increased, growth reductions were magnified and bacterial load was higher when both non-lethal stressors were combined. The combined exposure to the bacterium and predation risk considerably impaired the two key antipredator mechanisms of the damselfly larvae: they no longer reduced their food intake under predation risk and showed a synergistic reduction in escape swimming speed. The reinforcing negative effects on the fitness-related traits could be explained by the observed synergistic effects on food intake, swimming muscle mass, immune function and oxidative damage. These are likely widespread consequences of energetic constraints and increased metabolic rates associated with the fight-or-flight response. We therefore hypothesize that the here documented synergistic interactions with non-pathogenic bacteria may be widespread. Our results highlight the ignored ecological role of non-pathogenic bacteria in reinforcing the negative effects of predation risk on prey organisms.status: publishe

Chronic Predation Risk Reduces Escape Speed by Increasing Oxidative Damage: A Deadly Cost of an Adaptive Antipredator Response

Prey organisms evolved a multitude of plastic responses to avoid being eaten by predators. Besides the evolution of plastic morphological responses to escape predation, prey also evolved a set of physiological stress responses to avoid dying because of chronic predator stress per se due to disruption of cellular homeostasis. As physiological stress theory predicts increased energy consumption and the inhibition of essential nonemergency body functions, we tested whether chronic predation risk may increase oxidative damage thereby generating negative effects on escape performance. Specifically, we evaluated whether predation risk reduces escape swimming speed in damselfly larvae and whether this operates through stress-associated increases in oxidative damage. Counterintuitively and in contrast with many empirical studies, chronic predation risk decreased escape performance. This is however entirely consistent with the expectation of it being a long-term cost of responding to predation risk (e.g. by increasing respiration or upregulating the stress protein levels). The decreased swimming speed could be explained by an increased oxidative damage to proteins, thereby providing one of the poorly studied ecological links between oxidative damage and whole-animal performance. This likely widespread, understudied cost of chronic predation risk may provide an important pathway of non-consumptive predator effects on prey population dynamics. Moreover, it could play an evolutionary role by acting as a selective force causing prey organisms to adjust the magnitude of the physiological stress response and should be considered when evaluating life history trade-offs thought to be mediated by oxidative damage.status: publishe

Chlorpyrifos-induced oxidative damage is reduced under warming and predation risk: Explaining antagonistic interactions with a pesticide

Interactions with pollutants and environmental factors are poorly studied for physiological traits. Yet physiological traits are important for explaining and predicting interactions at higher levels of organization. We investigated the single and combined impact of the pesticide chlorpyrifos, predation risk and warming on endpoints related to oxidative stress in the damselfly Enallagma cyathigerum. We thereby integrated information on reactive oxygen species (ROS), antioxidant enzymes and oxidative damage. All three treatments impacted the oxidative stress levels and for most traits the pesticide interacted antagonistically with warming or predation risk. Chlorpyrifos exposure resulted in increased ROS levels, decreased antioxidant defence and increased oxidative damage compared to the control situation. Under warming, the pesticide-induced increase in oxidative stress was less strong and the investment in antioxidant defence higher. Although both the pesticide and predation risk increased oxidative damage, the effects of the pesticide on oxidative damage were less strong in the presence of predator cues (at 20 C). Despite the weaker pesticide-induced effects under predation risk, the combination of the pesticide and predator cues consistently caused the highest ROS levels, the lowest antioxidant defence and the highest oxidative damage, indicating the importance of cumulative stressor effects for impairing fitness. Our results provide the first evidence for antagonistic interactions of warming and predation risk with a pollutant for physiological traits. We identified two general mechanisms that may generate antagonistic interactions for oxidative stress: cross-tolerance and the maximum cumulative levels of damage.publisher: Elsevier articletitle: Chlorpyrifos-induced oxidative damage is reduced under warming and predation risk: Explaining antagonistic interactions with a pesticide journaltitle: Environmental Pollution articlelink: http://dx.doi.org/10.1016/j.envpol.2017.04.012 content_type: article copyright: © 2017 Elsevier Ltd. All rights reserved.status: publishe

Stronger effects of Roundup than its active ingredient glyphosate in damselfly larvae

Pesticides are causing strong decreases in aquatic biodiversity at concentrations assumed safe by legislation. One reason for the failing risk assessment may be strong differences in the toxicity of the active ingredient of pesticides and their commercial formulations. Sublethal effects, especially those on behaviour, have been largely ignored in this context, yet can be equally important as lethal effects at the population and ecosystem levels. Here, we compared the toxicity of the herbicide Roundup and its active ingredient glyphosate on survival, but also on ecologically relevant sublethal traits (life history, behaviour and physiology) in damselfly larvae. Roundup was more toxic than glyphosate with negative effects on survival, behaviour and most of the physiological traits being present at lower concentrations (food intake, escape swimming speed) or even only present (survival, sugar and total energy content and muscle mass) following Roundup exposure. This confirms the toxicity of the surfactant POEA. Notably, also glyphosate was not harmless: a realistic concentration of 2mg/l resulted in reduced growth rate, escape swimming speed and fat content. Our results therefore indicate that the toxicity of Roundup cannot be fully attributed to its surfactant, thereby suggesting that also the new generation of glyphosate-based herbicides with other mixtures of surfactants likely will have adverse effects on non-target aquatic organisms. Ecotoxicological studies comparing the toxicity of active ingredients and their commercial formulations typically ignore behaviour while the here observed differential effects on behaviour likely will negatively impact damselfly populations. Our data highlight that risk assessment of pesticides ignoring sublethal effects may contribute to the negative effects of pesticides on aquatic biodiversity.status: publishe

Predation risk causes oxidative damage in prey

While there is increasing interest in non-consumptive effects of predators on prey, physiological effects are understudied. While physiological stress responses play a crucial role in preparing escape responses, the increased metabolic rates and shunting of energy away from other body functions, including antioxidant defence, may generate costs in terms of increased oxidative stress. Here, we test whether predation risk increases oxidative damage in Enallagma cyathigerum damselfly larvae. Under predation risk, larvae showed higher lipid peroxidation, which was associated with lower levels of superoxide dismutase, a major antioxidant enzyme in insects, and higher superoxide anion concentrations, a potent reactive oxygen species. The mechanisms underlying oxidative damage are likely to be due to the shunting of energy away from antioxidant defence and to an increased metabolic rate, suggesting that the observed increased oxidative damage under predation risk may be widespread. Given the potentially severe fitness consequences of oxidative damage, this largely overlooked non-consumptive effect of predators may be contributing significantly to prey population dynamics.status: publishe

Fitness effects of chlorpyrifos in the damselfly Enallagma cyathigerum strongly depend upon temperature, food level and the fitness-related variable

Interactions between pollutants and suboptimal environmental conditions can have severe consequences for the toxicity of pollutants, yet are still poorly understood. To identify patterns across environmental conditions and across fitness-related variables we exposed Enallagma cyathigerum damselfly larvae to the pesticide chlorpyrifos at two food levels or at two temperatures and quantified four fitness-related variables (larval survival, development time, mass at emergence and adult cold resistance). Food level and temperature did not affect survival in the absence of the pesticide, yet the pesticide reduced survival only at the high temperature. Animals reacted to the pesticide by accelerating their development but only at the high food level and at the low temperature; at the low food level, however, pesticide exposure resulted in a slower development. Chlorpyrifos exposure resulted in smaller adults except in animals reared at the high food level. Animals reared at the low food level and at the low temperature had a higher cold resistance which was not affected by the pesticide. In summary our study highlight that combined effects of exposure to chlorpyrifos and the two environmental conditions (i) were mostly interactive and sometimes even reversed in comparison with the effect of the environmental condition in isolation, (ii) strongly differed depending on the fitness-related variable under study, (iii) were not always predictable based on the effect of the environmental condition in isolation, and (iv) bridged metamorphosis depending on which environmental condition was combined with the pesticide thereby potentially carrying over from aquatic to terrestrial ecosystems. These findings are relevant when extrapolating results of laboratory tests done under ideal environmental conditions to natural communities.status: publishe